Heretofore, it has been known to qualitatively determine the types of constituents in a sample to be analyzed by fractionating the sample using a chromatographic column. The fractions so obtained were then optically analyzed by scanning the fractions at various absorbance wavelengths, while being passed along the chromatographic column. A linear Beer's plot would then identify the type of substance in the sample.
Such a technique is described in "MOLECULAR SIEVE STUDIES OF INTERACTING PROTEIN SYSTEMS,"E. E. Brumbaugh and G. K. Ackers, Journal of Biological Chemistry, Vol. 243, No. 24, pp. 6315-6324, (1968) and
"MOLECULAR SIEVE STUDIES OF INTERATING PROTEIN SYSTEMS," E. E. Brumbaugh and G. K. Ackers, Analytical Biochemistry, Vol. 41, pp 543-559 (1971).
The present invention seeks to perform quantitative measurements with greater sensitivity of samples fractionated within a column by measuring the transmissivity of a light beam passing both through the column and one or more selected fractions of the sample.
When the prior art technique was used to quantitatively measure trace amounts of certain materials, it was discovered that the carrier fluid or the solvent passed through the chromotagraphic column increased the light transmissivity of the column. Such increased transmissivity was due to the difference in the respective indices of refraction of the fluid carrier and the chromotographic packing material. Such increased transmissivity, while small, would balance any decrease in transmissivity resulting from the trace amount of the constituent being analyzed. As a result, trace amounts of materials within a sample could not be measured, and, hence, the sensitivity of prior art systems was limited.
The present invention seeks to extend the sensitivity of chromatographic systems by nullifying (or compensating) the change in transmissivity due to the fluid carrier. Such compensation can be effected either optically or electronically.